Staphylococcus aureus is responsible for a number of human diseases, including skin and soft tissue infections. Annually, 292,000 hospitalizations in the US are due to S. aureus infections, of which 126,000 are related to methicillin-resistant Staphylococcus aureus (MRSA), resulting in 19,000 deaths. Enterococci are the leading cause of nosocomial bacteremia, surgical wound infections, and urinary tract infections. Vancomycin-resistant enterococci (VRE) accounts for >25% of the enterococci hospital-acquired infections in the US. A novel structural lead, the oxadiazol class of antibiotics, has emerged from our work. The oxadiazol antibiotics show high in vitro potency against Gram-positive bacteria comparable to those of linezolid and superior to vancomycin (both considered gold standards) and show in vivo activity. In addition, the oxadiazols have activity against vancomycin- resistant MRSA and VRE, two organisms for which treatment options are extremely limited. The compounds in hand at this point are not highly water soluble and their pharmacokinetic properties are not optimized. This project proposes to optimize the initial lead discovery to come up with lead candidates with the correct mix of pharmacological activity, PK attributes, and safety profile, such that an oxadiazol drug candidate will be selected and advanced to preclinical development. We also propose a novel method for the identification and validation of the target(s) for the oxadiazol antibiotics in the proteome of S. aureus and put forth a whole genome method for investigation of the mechanism of resistance to this class of antibiotics. Infections caused by drug-resistant bacteria are an increasing unmet medical need. The drug of last resort, linezolid, is reserved for use in hard-to-treat methicillin-resistant S. aureus (MRSA) and vancomycin-resistant enterococci (VRE). New treatments are urgently needed to combat the emergence of resistance to any antibiotic after introduction of the antibiotic to the clinic. Our group has developed a new oxadiazol class of antibiotics that shows activity against vancomycin-resistant MRSA and VRE comparable to linezolid. Our goal is to develop a new class of antibiotics to treat drug-resistant bacterial infections.